Three separate 5＇-flanking regions （1.1 kb, 1.9 kb and 2.2 kb） of crtO were cloned through walking upstream. Results of sequence analysis show that three 5＇-flanking regions of crtO might have some similar putati...Three separate 5＇-flanking regions （1.1 kb, 1.9 kb and 2.2 kb） of crtO were cloned through walking upstream. Results of sequence analysis show that three 5＇-flanking regions of crtO might have some similar putative cis-acting elements such as ABA （abscisic acid）-responsive element （ABRE）, C-repeat/dehydration responsive element （C-repeat/DRE）, light-responsive element （G-box, GAG-motif, I-box and ATC-motif）, wound-responsive element （WUN-motif）, auxin-responsive element （TGA-element）, MeJA-responsive element （TGACG-element） and MYB binding site （MBS）, except for typical TATA box or CCAAT box. These findings might mean diversiform regulatory patterns of crtO being in astaxanthin biosynthesis of Haematococcus pluvialis.展开更多
Objective: To analyze the genomic structure of SNC6, a progesterone\|receptor associated protein gene and its regulatory elements in its 5'\|flanking region. Methods: Genomic sequence from GenBank database (access...Objective: To analyze the genomic structure of SNC6, a progesterone\|receptor associated protein gene and its regulatory elements in its 5'\|flanking region. Methods: Genomic sequence from GenBank database (accession number: Z98048) covering the whole SNC6 gene was used to analyze the genomic structure of SNC6 and design primers for PCR amplification of its 5'\|flanking region. A 1894 bp fragment of the 5'\|flanking region \{(-1814\} to +75) was cloned by PCR using genomic DNA from a healthy donor peripheral blood lymphocyte as template. This fragment, as well as 3 shorter derivative fragments (1423 bp, 632 bp and 416 bp, which correspond to -1344 to +75, -552 to +75 and -337 to +75 respectively), were subcloned into pGL2 series luciferase reporter vectors. These constructs were introduced into colorectal cancer cell line SW620 for transient expression of reporter gene and luciferase activities were measured. Results: The genomic structure analysis showed there are 12 exons for SNC6 gene, which spans 32017 bp (nt71529 to nt39513 in Z98048 sequence). All transfected SW620 cells with the above 5\|flanking region\|containing constructs showed luciferase activities. The highest luciferase activities were measured in transfected cells with vectors containing 1894 bp fragments, and the lowest luciferase activities were measured in transfected cells with vectors containing 416 bp fragments. Luciferase activities were higher in transfected cells with vectors containing 632 bp fragments than that in transfected cells with vectors containing 1423 bp fragments. Conclusion: The basic transcription\|promoting element (promoter) for SNC6 expression resides between 0 to -337, and two transcription\|enhancing elements (enhancer) resides between -337 to -552 and -1344 to -1814, whereas one transcription\|inhibiting element (silencer) exists between -552 to -1344.展开更多
[Objective] The research aimed to construct the prokaryotic expression vector of VP5 protein of IBDV.The transmembrane region sequence of VP5 protein was knocked out.Moreover,the expression,separation and purification...[Objective] The research aimed to construct the prokaryotic expression vector of VP5 protein of IBDV.The transmembrane region sequence of VP5 protein was knocked out.Moreover,the expression,separation and purification of objective protein were carried out.[Method] PCR technology was used to respectively amplify the extracellular and intracellular fragments of VP5 gene of IBDV.Then,the two fragments were simultaneously linked to pET-28b（＋）,and it was the vector-intracellular fragment-extracellular fragment-vector.The recombinant expression plasmid pET-VP5-FC and the improved pET-VP5-SC of VP5 whose transmembrane region gene fragment was knocked out were constructed.Then,the expression plasmid was transformed into BL21（DE3）.After IPTG induction,the recombinant protein was purified by Ni affinity chromatography and the gel filtration chromatography.[Result] The soluble expressed VP5 of IBDV was obtained.[Conclusion] The research laid the foundation for further studying the structure and function of VP5 protein.展开更多
AIM: To evaluate the implication of substitutions in the hepatitis C virus (HCV) non-structural 5A (NS5A) protein in the resistance of HCV during mono-interferon (IFN) or combined IFN-ribavirin (IFN-R) therapy. Althou...AIM: To evaluate the implication of substitutions in the hepatitis C virus (HCV) non-structural 5A (NS5A) protein in the resistance of HCV during mono-interferon (IFN) or combined IFN-ribavirin (IFN-R) therapy. Although NS5A has been reported to interact with the HCV RNA- dependent RNA polymerase, NS5B, as well as with many cellular proteins, the function of NS5A in the life cycle of HCV remains unclear. METHODS: HCV quasispecies were studied by clon- ing and sequencing of sequential isolates from patients infected by HCV genotype 1b. Patients were treated by IFN-α2b for 3 mo followed by IFN-α2b alone or com- bined IFN-R therapy for 9 additional months. Patients were categorized intro two groups based on their re- sponse to the treatments: 7 with sustained virological re- sponse (SVR) (quasispecies = 150) and 3 non-respond- ers (NR) to IFN-R (quasispecies = 106). RESULTS: Prior to treatment, SVR patients displayed a lower complexity of quasispecies than NR patients. Most patients had a decrease in the complexity of quasispe- cies during therapy. Analysis of amino acids substitu- tions showed that the degree of the complexity of the interferon sensitivity-determining region (ISDR) and the V3 domain of NS5A protein was able to discriminate thetwo groups of patients. Moreover, SVR patients displayed more variability in the NS5A region than NR patients. CONCLUSION: These results suggest that detailed mo- lecular analysis of the NS5A region may be important for understanding its function in IFN response during HCV 1b infection.展开更多
The atmospheric water holding capacity will increase with temperature according to Clausius-Clapeyron scaling and affects precipitation.The rates of change in future precipitation extremes are quantified with changes ...The atmospheric water holding capacity will increase with temperature according to Clausius-Clapeyron scaling and affects precipitation.The rates of change in future precipitation extremes are quantified with changes in surface air temperature.Precipitation extremes in China are determined for the 21st century in six simulations using a regional climate model,RegCM4,and 17 global climate models that participated in CMIP5.First,we assess the performance of the CMIP5 models and RCM runs in their simulation of extreme precipitation for the current period(RF:1982-2001).The CMIP5 models and RCM results can capture the spatial variations of precipitation extremes,as well as those based on observations:OBS and XPP.Precipitation extremes over four subregions in China are predicted to increase in the mid-future(MF:2039-58)and far-future(FF:2079-98)relative to those for the RF period based on both the CMIP5 ensemble mean and RCM ensemble mean.The secular trends in the extremes of the CMIP5 models are predicted to increase from 2008 to 2058,and the RCM results show higher interannual variability relative to that of the CMIP5 models.Then,we quantify the increasing rates of change in precipitation extremes in the MF and FF periods in the subregions of China with the changes in surface air temperature.Finally,based on the water vapor equation,changes in precipitation extremes in China for the MF and FF periods are found to correlate positively with changes in the atmospheric vertical wind multiplied by changes in surface specific humidity(significant at the p<0.1 level).展开更多
Based on data of 22 models from the Coupled Model Inter-comparison Project Phase 5(CMIP5),the performance of climate simulation is assessed and future changes under RCP2.6,RCP4.5 and RCP8.5 are projected over critical...Based on data of 22 models from the Coupled Model Inter-comparison Project Phase 5(CMIP5),the performance of climate simulation is assessed and future changes under RCP2.6,RCP4.5 and RCP8.5 are projected over critical Belt and Road region.Compared with observations,the CMIP5 models simulate the linear trend and spatial distribution of the annual mean surface air temperature(SAT)better in the north(NBR)and south(SBR)of the Belt and Road region.The trend of the 22-model ensemble mean(CMIP5 MME)is 0.70/0.50 C per 100 years from 1901 to 2005,and the observed trend is 1.11/0.77 C per 100 years in the NBR/SBR region.After 1971,the relative error between CMIP5 MME and observations is 22%/15%in the NBR/SBR region.Seven/nine models are selected in the NBR/SBR to project future SAT changes under three RCP scenarios.For 2081e2100,warming in the NBR/SBR is projected to be(1.16±0.29)/(0.72±0.32)C,(2.41±0.54)/(1.55±0.44)C,and(5.23±1.02)/(3.33±0.65)C for RCP2.6,RCP4.5,and RCP8.5,respectively.Under the RCP scenarios,the NBR region shows greater warming than the SBR region.The most significant warming is expected in Kazakhstan and the northern part of the SBR.The associated uncertainty generally increases with time under the three RCP scenarios.Furthermore,increases in warming over the Belt and Road region are more remarkable under higher-emission scenarios than lower-emission ones.展开更多
Based on the outputs from 21 CMIP5 (Coupled Model Intercomparison Project phase 5) models, future changes in the mean temperature, precipitation and four climate extreme indices (annual maximum of daily maximum temper...Based on the outputs from 21 CMIP5 (Coupled Model Intercomparison Project phase 5) models, future changes in the mean temperature, precipitation and four climate extreme indices (annual maximum of daily maximum temperature (TXx), minimum of daily minimum temperature (TNn), annual total precipitation when the daily amount exceeds the 95th percentile of wet-day precipitation (R95p), and maximum consecutive 5-day precipitation (RX5day)) over Hindu Kush Himalayan (HKH) region are investigated under the greenhouse gas concentration pathways of RCP4.5 and RCP8.5. Two periods of the 21st century, 2036e2065 and 2066e2095, are selected, with the reference period is considered as 1976e2005. Results show general increase of the mean temperature, TXx and TNn under both scenarios, with the largest increases found during 2066e2095 under RCP8.5. Future precipitation is projected to increase over most part of HKH, except for the northwestern part. Intensification of the precipitation extremes is projected over the region. The uncertainties of mean temperature, TXx and TNn over the HKH1 subregions are the largest compared to the other three subregions and the overall HKH. Besides RX5day during 2036e2065 over HKH1, the uncertainties of R95p and RX5day tend to be larger following the increase of greenhouse gas concentrations. The multimodel ensemble medians of temperature and four extreme indices under RCP8.5 are projected to be larger than those under RCP4.5 in each of the subregions.展开更多
It is suggested that Δ6 fatty acid desaturase(FAD) plays a critical role in the biosynthesis of polyunsaturated fatty acids in plants and microalgae. But why does it adapt to the changed environments such as nitrogen...It is suggested that Δ6 fatty acid desaturase(FAD) plays a critical role in the biosynthesis of polyunsaturated fatty acids in plants and microalgae. But why does it adapt to the changed environments such as nitrogen starvation is seldom understood. One Δ6 FAD gene( MiD6 fad) from an arachidonic acidrich microalga M yrmecia incisa Reisigl(Chlorophyta) was first heterologously expressed in S accharomyces cerevisiae for the identification of function. The fatty acid profile of transgenic yeast detected by gas chromatography-mass spectrometry illustrated that the enzyme MiD6 FAD could convert linoleic and ?-linolenic acids to γ-linolenic and stearidonic acids, respectively, demonstrating that M iD6 fad encoded a Δ6 FAD. A 1 965-bp fragment of the cloned 2 347-bp 5′-upstream region of M iD6 fad was next subcloned and fused upstream with green fluorescent protein(GFP) gene to replace the GAL1 promoter of the vector pYES2. The generated construct was transformed into S. cerevisiae for function determination. Confocal microscopic images of the transformed line illustrated that this inserted fragment could drive GFP expression, which was further verified by fluorescence intensity quantification and Western blot analysis using antiGFP antibody. The conversion efficiency(approximately 2%-3%) of MiD6 FAD was much lower than the reported ? 3 FAD and Δ6 elongase in this microalga, suggesting that MiD6 FAD catalysed the possible ratelimiting step for ArA biosynthesis. The presence of several putative c is-acting regulatory elements in this identified promoter sheds new light on the regulation mechanism research of Δ6 FAD transcription for the ArA production in M. incisa in changing environmental factors.展开更多
Objectives: To obtain very end full-length cDNA ofhepatitis C virus (HCV) 5’ untranslated region(5’UTR) and analyze its primary and secondarystructure.Methods: A patient infected genotype 2a HCV wasidentified by rev...Objectives: To obtain very end full-length cDNA ofhepatitis C virus (HCV) 5’ untranslated region(5’UTR) and analyze its primary and secondarystructure.Methods: A patient infected genotype 2a HCV wasidentified by reverse transcription-nested polymerasechain reaction (RT-PCR) and restriction fragmentlength polymorphism (RFLP). Total RNA isolatedfrom the serum was used as template, and the cDNAof the 5’ untranslated region was amplified using rap-id amplification of cDNA ends (RACE). The frag-ments were recombinated by A-T clone strategy, andthe recombinants were confirmed by RFLP andPCR, and sequenced subsequently. Secondary struc-tures were analysed by RNAdraw.Results: Very end full-length cDNA of genotype 2aHCV 5’ UTR was obtained by RACE. In five clonesobtained, three contained full-length 5’UTR cDNA;A21G, G170A, T222C, T247C, C339T substitutionswere found as compared to HC-J6. Homological re-sults of HCV-1, HC-J6, HC-C2, HC-J8 were 93.6%-94.4%, 92.1%-93%, 98.8%-99.7%, 96.2%-96.5%, respectively; however, the substitutions didnot alter secondary structure. Two of 5 clones weredeletions of 53bp and 135bp at the 5’terminal ofHCV 5’UTR, respectively.Conclusions: RACE can be used to obtain the full-length cDNA of 2a genotype HCV 5’UTR. Genes de-leted at the 5’ terminal of HCV circulate in hepatitisC patients.展开更多
As an important part of gas hydrates, light hydrocarbons （LHs）, especially C5 to C7 hydrocarbons with various monomer compounds, provide a wide variety of geological and geochemical information, which have received ...As an important part of gas hydrates, light hydrocarbons （LHs）, especially C5 to C7 hydrocarbons with various monomer compounds, provide a wide variety of geological and geochemical information, which have received much attention from organic geochemists and petroleum geologists.展开更多
Objective:To study evolutionary relationship of the 5'untranslated regions(5'UTRs) in low passage dengue3 viruses(DEN3) isolated from hospitalized children with different clinical manifestations in Bangkok dur...Objective:To study evolutionary relationship of the 5'untranslated regions(5'UTRs) in low passage dengue3 viruses(DEN3) isolated from hospitalized children with different clinical manifestations in Bangkok during 24 year-evolution(1977-2000) comparing to the DEN3prototype(H87).Methods:The 5'UTRs of these Thai DEN3 and the H87 prototype were amplified by RT-PCR and sequenced.Their multiple sequence alignments were done by Codon Code Aligner v 4.0.4 software and their RNA secondary structures were predicted by MFOLD software.Replication of five Thai DEN3 candidates comparing to the 1187 prototype were done in human(HepG2) and the mosquito(C6/36) cell lines.Results:Among these Thai DEN3,the completely identical sequences of their first 89 nucleotides,their high-order secondary structure of 5'UTRs and three SNPs including the predominant C90 T,and two minor SNPs including A109 G and A112 G were found.The C90 T of Thai DEN3.Bangkok isolates was shown predominantly before 1977.Five Thai DEN3 candidates with the predominant C90 T were shown to replicate in human(HepG2) and the mosquito(C6/36) cell lines better than the H87 prototype.However,their highly conserved sequences as well as SNPs of the 5'UTR did not appear to correlate with their disease severity in human.Conclusions:Our findings highlighted evolutionary relationship of the completely identical 89 nucleotide sequence,the high-order secondary structure and the predominant C90 T of the 5'UTR of these Thai DEN3 during 24 year-evolution further suggesting to be their genetic markers and magic targets for future research on antiviral therapy as well as vaccine approaches of Thai DEN3.展开更多
The aim of this study was to determine the effects of an IL-6 gene polymorphism, discovered in the 5＇ regulatory region, on porcine litter size. An association analysis was performed between the polymorphism and tota...The aim of this study was to determine the effects of an IL-6 gene polymorphism, discovered in the 5＇ regulatory region, on porcine litter size. An association analysis was performed between the polymorphism and total number born （TNB） and number born alive （NBA） in 421 sows. The polymorphism was at Hpy188I within the 5＇ regulatory region of IL- 6 gene. Three genotypes of AA, AG, and GG were detected in Landrace, and two genotypes, AA and AG, were detected in Yorkshire and Duroc pigs. The A allele was the superior allele in all three breeds, with allele frequencies ranging from 0. 901 to 0.993. The IL-6 genotype was highly significantly associated with TNB and NBA in the third and following parities （ P 〈 0.01 ）, and with total parities （ P 〈 0.05）. In general, the TNB and NBA showed a tendency of GG 〉 AG 〉 AA, indicating that the common allele was the least favorable for litter size. Thus, there is an enormous opportunity to increase litter size if this effect is confirmed in other studies.展开更多
AIM: To investigate the mutations of the 5' noncoding region of BCL-6 gene in Chinese patients with primary gastric lymphomas. METHODS: PCR and direct DNA sequencing were used to identify BCL-6 gene mutations in t...AIM: To investigate the mutations of the 5' noncoding region of BCL-6 gene in Chinese patients with primary gastric lymphomas. METHODS: PCR and direct DNA sequencing were used to identify BCL-6 gene mutations in the 5' noncoding region in 29 cases of gastric diffuse large B-cell lymphoma (DLBCL) and 18 cases of gastric mucosa-associated lymphoid tissue (MALT) lymphoma as well as 10 cases of reactive hyperplasia of lymph node (LRH). RESULTS: Six of 29 gastric DLBCLs (20.7%), 4 of 18 gastric MALT lymphomas (22.2%) and 1 of 10 LRHs(10%) were found to have mutations. All mutations were single-base substitutions and the frequency of single-base changes was 0.20×1O^(-2)-1.02×1O^(-2)per bp. CONCLUSION: Point mutations in the 5' noncoding region of BCL-6 gene are found in Chinese patients with primary gastric DLBCLs and MALT lymphomas, suggesting that they may, in some extent, participate in the pathogenesis of primary gastric DLBCLs and MALT lymphomas.展开更多
To determine the number of mutations in the NS5A region of the hepatitis C virus (HCV) and its relationship to the response to antiviral therapy in patients with chronic hepatitis C genotype 1 who are non-responders t...To determine the number of mutations in the NS5A region of the hepatitis C virus (HCV) and its relationship to the response to antiviral therapy in patients with chronic hepatitis C genotype 1 who are non-responders to two or more treatments. METHODSSequences within HCV NS5A [PKR binding domain (PKRBD) and the interferon-sensitivity-determining region (ISDR)] were analysed via direct sequencing in a selected cohort of 72 patients, with a total of 201 treatments [interferon-alpha (IFN-α), n = 49; IFN-α + ribavirin (RBV), n = 75; pegylated (peg) IFN-α + RBV, n = 47; first-generation direct-acting antivirals (DAAs), n = 13; and second-generation DAAs, n = 17]. Of these, 48/201 achieved a sustained virological response (SVR) and 153/201 achieved no virological response (NVR). RESULTSFor both regions, treatments resulting in SVR were associated with more baseline mutations than were treatments resulting in NVR (SVR vs NVR; PKRBD: 5.82 ± 3 vs 4.86 ± 2 mutations, P = 0.045; ISDR: 2.65 ± 2 vs 1.51 ± 1.7 mutations, P = 0.005). A decrease or no change in the number of mutations over time between treatments in the PKRBD or ISDR, as shown by sequencing, was associated with patients who usually failed to respond to treatment (PKRBD, P = 0.02; ISDR, P = 0.001). Moreover, patients showing a post-treatment baseline viral load > 600000 IU/mL and increased ISDR mutations with respect to the previous treatment were 9.21 times more likely to achieve SVR (P = 0.001). CONCLUSIONThe obtained results show that among patients who have shown no response to two or more antiviral treatments, the likelihood of achieving SVR increases with the genetic variability in the ISDR region (≥ 2 mutations or number of substitutions from the HCV-J and HCV-1 prototype), especially when the viral load is greater than 600000 IU/mL.展开更多
Dear Editor： Increased homocysteine levels due to vitamin B6 or B12 deficiency or genetic defects in folate pathway genes are associated with an increased incidence of non-syndromic cleft lip with or without cleft p...Dear Editor： Increased homocysteine levels due to vitamin B6 or B12 deficiency or genetic defects in folate pathway genes are associated with an increased incidence of non-syndromic cleft lip with or without cleft palate （NSCLP）tlj. Thymidylate synthase （TS） is a folate-dependent enzyme that catalyzes methylation of 2＇-deoxyuridine-5＇-monophosphate （dUMP） to 2＇-deox- ythymidine-5＇-monophosphate （dTMP）, a rate-limiting step in DNA synthesis,展开更多
Through the analysis of ensembles of coupled model simulations and projections collected from CMIP3 and CMIP5, we demonstrate that a fundamental spatial scale limit might exist below which useful additional refinement...Through the analysis of ensembles of coupled model simulations and projections collected from CMIP3 and CMIP5, we demonstrate that a fundamental spatial scale limit might exist below which useful additional refinement of climate model predictions and projections may not be possible. That limit varies among climate variables and from region to region. We show that the uncertainty（noise） in surface temperature predictions（represented by the spread among an ensemble of global climate model simulations） generally exceeds the ensemble mean（signal） at horizontal scales below 1000 km throughout North America, implying poor predictability at those scales. More limited skill is shown for the predictability of regional precipitation. The ensemble spread in this case tends to exceed or equal the ensemble mean for scales below 2000 km. These findings highlight the challenges in predicting regionally specific future climate anomalies, especially for hydroclimatic impacts such as drought and wetness.展开更多
基金supported by the National Natural Science Foundation (40706050 40706048)+4 种基金Natural Science Foundation of Shandong University of Technology (4040306017)Start-up Foundation for Ph.D of Shandong University of Technology (4041-4050174041-405016)National Key Technology R&D program(11200602)Foundation of State Commonweal Institute (2060402/2)
文摘Three separate 5＇-flanking regions （1.1 kb, 1.9 kb and 2.2 kb） of crtO were cloned through walking upstream. Results of sequence analysis show that three 5＇-flanking regions of crtO might have some similar putative cis-acting elements such as ABA （abscisic acid）-responsive element （ABRE）, C-repeat/dehydration responsive element （C-repeat/DRE）, light-responsive element （G-box, GAG-motif, I-box and ATC-motif）, wound-responsive element （WUN-motif）, auxin-responsive element （TGA-element）, MeJA-responsive element （TGACG-element） and MYB binding site （MBS）, except for typical TATA box or CCAAT box. These findings might mean diversiform regulatory patterns of crtO being in astaxanthin biosynthesis of Haematococcus pluvialis.
文摘Objective: To analyze the genomic structure of SNC6, a progesterone\|receptor associated protein gene and its regulatory elements in its 5'\|flanking region. Methods: Genomic sequence from GenBank database (accession number: Z98048) covering the whole SNC6 gene was used to analyze the genomic structure of SNC6 and design primers for PCR amplification of its 5'\|flanking region. A 1894 bp fragment of the 5'\|flanking region \{(-1814\} to +75) was cloned by PCR using genomic DNA from a healthy donor peripheral blood lymphocyte as template. This fragment, as well as 3 shorter derivative fragments (1423 bp, 632 bp and 416 bp, which correspond to -1344 to +75, -552 to +75 and -337 to +75 respectively), were subcloned into pGL2 series luciferase reporter vectors. These constructs were introduced into colorectal cancer cell line SW620 for transient expression of reporter gene and luciferase activities were measured. Results: The genomic structure analysis showed there are 12 exons for SNC6 gene, which spans 32017 bp (nt71529 to nt39513 in Z98048 sequence). All transfected SW620 cells with the above 5\|flanking region\|containing constructs showed luciferase activities. The highest luciferase activities were measured in transfected cells with vectors containing 1894 bp fragments, and the lowest luciferase activities were measured in transfected cells with vectors containing 416 bp fragments. Luciferase activities were higher in transfected cells with vectors containing 632 bp fragments than that in transfected cells with vectors containing 1423 bp fragments. Conclusion: The basic transcription\|promoting element (promoter) for SNC6 expression resides between 0 to -337, and two transcription\|enhancing elements (enhancer) resides between -337 to -552 and -1344 to -1814, whereas one transcription\|inhibiting element (silencer) exists between -552 to -1344.
基金Supported by the National Natural Science Fundation Item of China(30970578,31070651)"Excellent Talent Support Plan in NewCentury"of Ministry of Education(NECT-08-0731)~~
文摘[Objective] The research aimed to construct the prokaryotic expression vector of VP5 protein of IBDV.The transmembrane region sequence of VP5 protein was knocked out.Moreover,the expression,separation and purification of objective protein were carried out.[Method] PCR technology was used to respectively amplify the extracellular and intracellular fragments of VP5 gene of IBDV.Then,the two fragments were simultaneously linked to pET-28b（＋）,and it was the vector-intracellular fragment-extracellular fragment-vector.The recombinant expression plasmid pET-VP5-FC and the improved pET-VP5-SC of VP5 whose transmembrane region gene fragment was knocked out were constructed.Then,the expression plasmid was transformed into BL21（DE3）.After IPTG induction,the recombinant protein was purified by Ni affinity chromatography and the gel filtration chromatography.[Result] The soluble expressed VP5 of IBDV was obtained.[Conclusion] The research laid the foundation for further studying the structure and function of VP5 protein.
基金Supported by a grant from l’Agence National de la Recherche sur le Sida (ANRS grant 2001/011)
文摘AIM: To evaluate the implication of substitutions in the hepatitis C virus (HCV) non-structural 5A (NS5A) protein in the resistance of HCV during mono-interferon (IFN) or combined IFN-ribavirin (IFN-R) therapy. Although NS5A has been reported to interact with the HCV RNA- dependent RNA polymerase, NS5B, as well as with many cellular proteins, the function of NS5A in the life cycle of HCV remains unclear. METHODS: HCV quasispecies were studied by clon- ing and sequencing of sequential isolates from patients infected by HCV genotype 1b. Patients were treated by IFN-α2b for 3 mo followed by IFN-α2b alone or com- bined IFN-R therapy for 9 additional months. Patients were categorized intro two groups based on their re- sponse to the treatments: 7 with sustained virological re- sponse (SVR) (quasispecies = 150) and 3 non-respond- ers (NR) to IFN-R (quasispecies = 106). RESULTS: Prior to treatment, SVR patients displayed a lower complexity of quasispecies than NR patients. Most patients had a decrease in the complexity of quasispe- cies during therapy. Analysis of amino acids substitu- tions showed that the degree of the complexity of the interferon sensitivity-determining region (ISDR) and the V3 domain of NS5A protein was able to discriminate thetwo groups of patients. Moreover, SVR patients displayed more variability in the NS5A region than NR patients. CONCLUSION: These results suggest that detailed mo- lecular analysis of the NS5A region may be important for understanding its function in IFN response during HCV 1b infection.
基金`This study was supported by the National Key Research and Development Program of China(Grant No.2019YFA0606903)the National Natural Science Foundation of China(Grant No.42075162)the Strategic Priority Research Program of Chinese Academy of Sciences(Grant No.XDA23090102).
文摘The atmospheric water holding capacity will increase with temperature according to Clausius-Clapeyron scaling and affects precipitation.The rates of change in future precipitation extremes are quantified with changes in surface air temperature.Precipitation extremes in China are determined for the 21st century in six simulations using a regional climate model,RegCM4,and 17 global climate models that participated in CMIP5.First,we assess the performance of the CMIP5 models and RCM runs in their simulation of extreme precipitation for the current period(RF:1982-2001).The CMIP5 models and RCM results can capture the spatial variations of precipitation extremes,as well as those based on observations:OBS and XPP.Precipitation extremes over four subregions in China are predicted to increase in the mid-future(MF:2039-58)and far-future(FF:2079-98)relative to those for the RF period based on both the CMIP5 ensemble mean and RCM ensemble mean.The secular trends in the extremes of the CMIP5 models are predicted to increase from 2008 to 2058,and the RCM results show higher interannual variability relative to that of the CMIP5 models.Then,we quantify the increasing rates of change in precipitation extremes in the MF and FF periods in the subregions of China with the changes in surface air temperature.Finally,based on the water vapor equation,changes in precipitation extremes in China for the MF and FF periods are found to correlate positively with changes in the atmospheric vertical wind multiplied by changes in surface specific humidity(significant at the p<0.1 level).
基金This work is founded by the National Key Research and Development Program of China(2016YFA0602703 and 2016YFA0600704),and the National Natural Science Foundation of China(41330527).
文摘Based on data of 22 models from the Coupled Model Inter-comparison Project Phase 5(CMIP5),the performance of climate simulation is assessed and future changes under RCP2.6,RCP4.5 and RCP8.5 are projected over critical Belt and Road region.Compared with observations,the CMIP5 models simulate the linear trend and spatial distribution of the annual mean surface air temperature(SAT)better in the north(NBR)and south(SBR)of the Belt and Road region.The trend of the 22-model ensemble mean(CMIP5 MME)is 0.70/0.50 C per 100 years from 1901 to 2005,and the observed trend is 1.11/0.77 C per 100 years in the NBR/SBR region.After 1971,the relative error between CMIP5 MME and observations is 22%/15%in the NBR/SBR region.Seven/nine models are selected in the NBR/SBR to project future SAT changes under three RCP scenarios.For 2081e2100,warming in the NBR/SBR is projected to be(1.16±0.29)/(0.72±0.32)C,(2.41±0.54)/(1.55±0.44)C,and(5.23±1.02)/(3.33±0.65)C for RCP2.6,RCP4.5,and RCP8.5,respectively.Under the RCP scenarios,the NBR region shows greater warming than the SBR region.The most significant warming is expected in Kazakhstan and the northern part of the SBR.The associated uncertainty generally increases with time under the three RCP scenarios.Furthermore,increases in warming over the Belt and Road region are more remarkable under higher-emission scenarios than lower-emission ones.
文摘Based on the outputs from 21 CMIP5 (Coupled Model Intercomparison Project phase 5) models, future changes in the mean temperature, precipitation and four climate extreme indices (annual maximum of daily maximum temperature (TXx), minimum of daily minimum temperature (TNn), annual total precipitation when the daily amount exceeds the 95th percentile of wet-day precipitation (R95p), and maximum consecutive 5-day precipitation (RX5day)) over Hindu Kush Himalayan (HKH) region are investigated under the greenhouse gas concentration pathways of RCP4.5 and RCP8.5. Two periods of the 21st century, 2036e2065 and 2066e2095, are selected, with the reference period is considered as 1976e2005. Results show general increase of the mean temperature, TXx and TNn under both scenarios, with the largest increases found during 2066e2095 under RCP8.5. Future precipitation is projected to increase over most part of HKH, except for the northwestern part. Intensification of the precipitation extremes is projected over the region. The uncertainties of mean temperature, TXx and TNn over the HKH1 subregions are the largest compared to the other three subregions and the overall HKH. Besides RX5day during 2036e2065 over HKH1, the uncertainties of R95p and RX5day tend to be larger following the increase of greenhouse gas concentrations. The multimodel ensemble medians of temperature and four extreme indices under RCP8.5 are projected to be larger than those under RCP4.5 in each of the subregions.
基金Supported by the National Natural Science Foundation of China(No.31172389)the Special Project of Marine Renewable Energy from the State Oceanic Administration(No.SHME2011SW02)the Shanghai Universities Peak Discipline Project of Aquaculture
文摘It is suggested that Δ6 fatty acid desaturase(FAD) plays a critical role in the biosynthesis of polyunsaturated fatty acids in plants and microalgae. But why does it adapt to the changed environments such as nitrogen starvation is seldom understood. One Δ6 FAD gene( MiD6 fad) from an arachidonic acidrich microalga M yrmecia incisa Reisigl(Chlorophyta) was first heterologously expressed in S accharomyces cerevisiae for the identification of function. The fatty acid profile of transgenic yeast detected by gas chromatography-mass spectrometry illustrated that the enzyme MiD6 FAD could convert linoleic and ?-linolenic acids to γ-linolenic and stearidonic acids, respectively, demonstrating that M iD6 fad encoded a Δ6 FAD. A 1 965-bp fragment of the cloned 2 347-bp 5′-upstream region of M iD6 fad was next subcloned and fused upstream with green fluorescent protein(GFP) gene to replace the GAL1 promoter of the vector pYES2. The generated construct was transformed into S. cerevisiae for function determination. Confocal microscopic images of the transformed line illustrated that this inserted fragment could drive GFP expression, which was further verified by fluorescence intensity quantification and Western blot analysis using antiGFP antibody. The conversion efficiency(approximately 2%-3%) of MiD6 FAD was much lower than the reported ? 3 FAD and Δ6 elongase in this microalga, suggesting that MiD6 FAD catalysed the possible ratelimiting step for ArA biosynthesis. The presence of several putative c is-acting regulatory elements in this identified promoter sheds new light on the regulation mechanism research of Δ6 FAD transcription for the ArA production in M. incisa in changing environmental factors.
基金This work was supported by two grants from National Science Foundation of China (No: 39770684, 30170844).
文摘Objectives: To obtain very end full-length cDNA ofhepatitis C virus (HCV) 5’ untranslated region(5’UTR) and analyze its primary and secondarystructure.Methods: A patient infected genotype 2a HCV wasidentified by reverse transcription-nested polymerasechain reaction (RT-PCR) and restriction fragmentlength polymorphism (RFLP). Total RNA isolatedfrom the serum was used as template, and the cDNAof the 5’ untranslated region was amplified using rap-id amplification of cDNA ends (RACE). The frag-ments were recombinated by A-T clone strategy, andthe recombinants were confirmed by RFLP andPCR, and sequenced subsequently. Secondary struc-tures were analysed by RNAdraw.Results: Very end full-length cDNA of genotype 2aHCV 5’ UTR was obtained by RACE. In five clonesobtained, three contained full-length 5’UTR cDNA;A21G, G170A, T222C, T247C, C339T substitutionswere found as compared to HC-J6. Homological re-sults of HCV-1, HC-J6, HC-C2, HC-J8 were 93.6%-94.4%, 92.1%-93%, 98.8%-99.7%, 96.2%-96.5%, respectively; however, the substitutions didnot alter secondary structure. Two of 5 clones weredeletions of 53bp and 135bp at the 5’terminal ofHCV 5’UTR, respectively.Conclusions: RACE can be used to obtain the full-length cDNA of 2a genotype HCV 5’UTR. Genes de-leted at the 5’ terminal of HCV circulate in hepatitisC patients.
基金financially supported by the National Natural Science Foundation of China(grant No.41273066)
文摘As an important part of gas hydrates, light hydrocarbons （LHs）, especially C5 to C7 hydrocarbons with various monomer compounds, provide a wide variety of geological and geochemical information, which have received much attention from organic geochemists and petroleum geologists.
基金supported by two research grants of Associate Professor Dr.W.Attatippaholkun:Grant No.493-5600-G-00-3461,Program in Science and Technology Cooperation,Human Capacity Development,Bureau for Global Programs,Field Support and Research,US Agency for International Development,Washington,DCThe Royal Golden Jubilee-Ph.D Program,Thailand Research Fund,Thailand
文摘Objective:To study evolutionary relationship of the 5'untranslated regions(5'UTRs) in low passage dengue3 viruses(DEN3) isolated from hospitalized children with different clinical manifestations in Bangkok during 24 year-evolution(1977-2000) comparing to the DEN3prototype(H87).Methods:The 5'UTRs of these Thai DEN3 and the H87 prototype were amplified by RT-PCR and sequenced.Their multiple sequence alignments were done by Codon Code Aligner v 4.0.4 software and their RNA secondary structures were predicted by MFOLD software.Replication of five Thai DEN3 candidates comparing to the 1187 prototype were done in human(HepG2) and the mosquito(C6/36) cell lines.Results:Among these Thai DEN3,the completely identical sequences of their first 89 nucleotides,their high-order secondary structure of 5'UTRs and three SNPs including the predominant C90 T,and two minor SNPs including A109 G and A112 G were found.The C90 T of Thai DEN3.Bangkok isolates was shown predominantly before 1977.Five Thai DEN3 candidates with the predominant C90 T were shown to replicate in human(HepG2) and the mosquito(C6/36) cell lines better than the H87 prototype.However,their highly conserved sequences as well as SNPs of the 5'UTR did not appear to correlate with their disease severity in human.Conclusions:Our findings highlighted evolutionary relationship of the completely identical 89 nucleotide sequence,the high-order secondary structure and the predominant C90 T of the 5'UTR of these Thai DEN3 during 24 year-evolution further suggesting to be their genetic markers and magic targets for future research on antiviral therapy as well as vaccine approaches of Thai DEN3.
基金supported by National Natural Science Foundation of China ( No.31172176 )China Agriculture Research System ( No.CARS-36 )
文摘The aim of this study was to determine the effects of an IL-6 gene polymorphism, discovered in the 5＇ regulatory region, on porcine litter size. An association analysis was performed between the polymorphism and total number born （TNB） and number born alive （NBA） in 421 sows. The polymorphism was at Hpy188I within the 5＇ regulatory region of IL- 6 gene. Three genotypes of AA, AG, and GG were detected in Landrace, and two genotypes, AA and AG, were detected in Yorkshire and Duroc pigs. The A allele was the superior allele in all three breeds, with allele frequencies ranging from 0. 901 to 0.993. The IL-6 genotype was highly significantly associated with TNB and NBA in the third and following parities （ P 〈 0.01 ）, and with total parities （ P 〈 0.05）. In general, the TNB and NBA showed a tendency of GG 〉 AG 〉 AA, indicating that the common allele was the least favorable for litter size. Thus, there is an enormous opportunity to increase litter size if this effect is confirmed in other studies.
基金Supported by National Natural Science Foundation of China,No.39900065
文摘AIM: To investigate the mutations of the 5' noncoding region of BCL-6 gene in Chinese patients with primary gastric lymphomas. METHODS: PCR and direct DNA sequencing were used to identify BCL-6 gene mutations in the 5' noncoding region in 29 cases of gastric diffuse large B-cell lymphoma (DLBCL) and 18 cases of gastric mucosa-associated lymphoid tissue (MALT) lymphoma as well as 10 cases of reactive hyperplasia of lymph node (LRH). RESULTS: Six of 29 gastric DLBCLs (20.7%), 4 of 18 gastric MALT lymphomas (22.2%) and 1 of 10 LRHs(10%) were found to have mutations. All mutations were single-base substitutions and the frequency of single-base changes was 0.20×1O^(-2)-1.02×1O^(-2)per bp. CONCLUSION: Point mutations in the 5' noncoding region of BCL-6 gene are found in Chinese patients with primary gastric DLBCLs and MALT lymphomas, suggesting that they may, in some extent, participate in the pathogenesis of primary gastric DLBCLs and MALT lymphomas.
基金Supported by“Consejería de Salud de la Junta de Andalucía”,No.PI0137/07“Instituto de Salud CarlosⅢ”,No.FISIntrasalud PI010/717
文摘To determine the number of mutations in the NS5A region of the hepatitis C virus (HCV) and its relationship to the response to antiviral therapy in patients with chronic hepatitis C genotype 1 who are non-responders to two or more treatments. METHODSSequences within HCV NS5A [PKR binding domain (PKRBD) and the interferon-sensitivity-determining region (ISDR)] were analysed via direct sequencing in a selected cohort of 72 patients, with a total of 201 treatments [interferon-alpha (IFN-α), n = 49; IFN-α + ribavirin (RBV), n = 75; pegylated (peg) IFN-α + RBV, n = 47; first-generation direct-acting antivirals (DAAs), n = 13; and second-generation DAAs, n = 17]. Of these, 48/201 achieved a sustained virological response (SVR) and 153/201 achieved no virological response (NVR). RESULTSFor both regions, treatments resulting in SVR were associated with more baseline mutations than were treatments resulting in NVR (SVR vs NVR; PKRBD: 5.82 ± 3 vs 4.86 ± 2 mutations, P = 0.045; ISDR: 2.65 ± 2 vs 1.51 ± 1.7 mutations, P = 0.005). A decrease or no change in the number of mutations over time between treatments in the PKRBD or ISDR, as shown by sequencing, was associated with patients who usually failed to respond to treatment (PKRBD, P = 0.02; ISDR, P = 0.001). Moreover, patients showing a post-treatment baseline viral load > 600000 IU/mL and increased ISDR mutations with respect to the previous treatment were 9.21 times more likely to achieve SVR (P = 0.001). CONCLUSIONThe obtained results show that among patients who have shown no response to two or more antiviral treatments, the likelihood of achieving SVR increases with the genetic variability in the ISDR region (≥ 2 mutations or number of substitutions from the HCV-J and HCV-1 prototype), especially when the viral load is greater than 600000 IU/mL.
基金funding from the Indian Council of Medical Research(ICMR),Government of India(Project Ref.No.56/15/2007-BMS)
文摘Dear Editor： Increased homocysteine levels due to vitamin B6 or B12 deficiency or genetic defects in folate pathway genes are associated with an increased incidence of non-syndromic cleft lip with or without cleft palate （NSCLP）tlj. Thymidylate synthase （TS） is a folate-dependent enzyme that catalyzes methylation of 2＇-deoxyuridine-5＇-monophosphate （dUMP） to 2＇-deox- ythymidine-5＇-monophosphate （dTMP）, a rate-limiting step in DNA synthesis,
基金partially supported by the NSF(Grant No.AGS-1305798)the ONR(Grant No.N000140910526)
文摘Through the analysis of ensembles of coupled model simulations and projections collected from CMIP3 and CMIP5, we demonstrate that a fundamental spatial scale limit might exist below which useful additional refinement of climate model predictions and projections may not be possible. That limit varies among climate variables and from region to region. We show that the uncertainty（noise） in surface temperature predictions（represented by the spread among an ensemble of global climate model simulations） generally exceeds the ensemble mean（signal） at horizontal scales below 1000 km throughout North America, implying poor predictability at those scales. More limited skill is shown for the predictability of regional precipitation. The ensemble spread in this case tends to exceed or equal the ensemble mean for scales below 2000 km. These findings highlight the challenges in predicting regionally specific future climate anomalies, especially for hydroclimatic impacts such as drought and wetness.
